The maxillary canal of the titanosuchid Jonkeria (Synapsida, Dinocephalia)

The maxillary canal of the titanosuchid dinocephalian Jonkeria is described based on digitised serial sections. We highlight that its morphology is more like that of the tapinocephalid Moschognathus than that of Anteosaurus. This is unexpected given the similarities between the dentition of Jonkeria and Anteosaurus (i.e., presence of a canine) and the fact that the branching pattern of the maxillary canal in synapsids usually co-varies with dentition. Hypotheses to account for similarities between Jonkeria and Moschognathus (common ancestry, function in social signalling or underwater sensing) are discussed. It is likely that the maxillary canal carries a strong phylogenetic signal, here supporting the clade Tapinocephalia. Supplementary Information The online version contains supplementary material available at 10.1007/s00114-023-01853-w.


Photography
Sections were photographed using a Canon EOS 450D fitted with a Canon EF-S 15-85 mm f/3.5-5.6 IS USM lens mounted to a tripod. Image stabilisation (IS) was disabled, and a hot-shoe mounted spirit level was used to ensure that the image sensor was level. A remote shutter release (Canon RS-60E3) was used to actuate the shutter without the need to handle the camera set up between photographs. The working distance between image sensor and specimen was kept as consistent as possible; however slight variation may have occurred due to differences in slice thickness. For all photographs the ISO and aperture were set at 100 and f/16, respectively.

Image Processing
The raw JPG images (4272 × 2848 pixels) were imported into GIMP (https://gimp.org) for processing (e.g., Fig.   S1a-c). Due to the nature of the sections, each alternate photograph needed to be flipped/mirrored for the anatomical structures to be in the same orientation (e.g., Fig. S1b). Some images needed to be rotated 90° as Page 2 of 4 they were photographed in a different orientation. The resulting image stack was cropped to reduce the file sizes. A black background layer was inserted at the bottom of the image stack so that after rotating and cropping, the resulting images would have a consistent image size of 3035 × 3181 pixels. In addition, a black mask was applied to the area surrounding the specimen in order to removed excess 'noise' (e.g., accumulated dust and cloth background). Note that noise removal is optional, but can improve the results of automated 'thresholding' in the later steps (e.g., segmentation). No other processing (e.g., colour correction, image sharpening, etc.) was applied to the images. The processed image stack was exported as 61 individual PNG files (e.g., Figs S1d-f).

Image Alignment
The PNG stack was loaded into SPIERSalign V 3.1.0 (Sutton et al. 2012; https://spiers-software.org). Fiduciary markers were set on the straight edges of the middle image (Slice 31; Fig. S2). Alignment was performed manually, using the 'Rotate' and 'Shift' commands from the Transform dropdown menu. A 'first pass' through the image stack ensured that the region of interest for all 61 images would remain within the 3035 × 3181 pixel limit (orange vertical markers in Fig. S2). A 'second pass' was then undertaken to refine the rotation of the images, aligning them first to the blue horizontal marker, and then to the orange vertical marker on the right (Fig. S2). The stack of aligned images was cropped in SPIERSalign, and the resulting images were exported as TIF files.

Reconstruction
The TIF stack was loaded into 3D Slicer (Kikinis et al. 2014; https://slicer.org) using the 'ImageStacks' function within the SlicerMorph extension (Rolfe et al. 2021; https://slicermorph.github.io). We are unsure of the thickness of the disc used to create the sections. A modest estimate of 4 mm was used, thus the volume was reconstructed with a slice interval of 6 mm. The reconstructed volume was exported as an NRRD file.

Segmentation
The maxillary canal and teeth were manually segmented using the commercially available software Avizo 9 (Thermo Fisher Scientific, Hillsborough, OR, USA). Similar results could be obtained from using the SPIERSedit module, or other 3-D visualisation software (see Buser et al. 2020). Screenshots of the final segmented data were exported as TIF images (Fig. 1).

Comment on 'Resolution' of Reconstructions
The focus of this study was to describe the gross morphology of the maxillary canals in a serially sectioned specimen of Jonkeria truculenta, with particular interest in the positions of the various branches of these canals relative to other identifiable anatomical structures (e.g., canine alveolus, etc.). Although the reconstructed slice interval of 6 mm is of a relatively low resolution by modern standards (cf. 0.102 mm voxel size achieved from X-ray microtomography (Benoit et al. 2021)). However, the large size of these animals (skull length of Jonkeria ~520-680 mm (Jirah 2022)), means that this slice interval is adequate for recording sufficient morphology to make the description of the maxillary canals feasible. As such, the 'jagged' appearance of the raw 3-D reconstruction has a negligible effect on the descriptions in the main text.